犏牛PPP1R11基因的克隆及在睾丸中的表达规律研究

旨在克隆犏牛蛋白磷酸酶1调节亚基11(protein phosphatase 1 regulatory subunit 11,PPP1R11)基因,分析其在不同发育阶段睾丸中的表达与定位,为解析其在雄性生殖中的功能机制提供理论依据.本研究采集成年犏牛睾丸、附睾、心、肝、脾、肺、肾、大肠、小肠、胃、肌肉和脂肪组织(n=3...

Cloning and Expression Analysis of Cattle-yak PPP1R11 Gene in Testis

This study was conducted to clone the PPP1R11 gene in cattle-yak,detect the expression and location in testes of cattle-yak at different developmental stages,as to provide a theoretical basis for further research of the mechanism of function in male reproduction.The testis,epididymis,heart,liver,spleen,lung,kidney,large intestine,small intestine,stomach,muscle and adipose tissue of adult cattle-yak were collected(n=3),testicles of cattle-yak and yak in the fetal period(5-6 months old),juvenile period(1-2 years old)and adult period(3-4 years old)(n=3)were collected as experimental materials.The PPP1R11 CDS region of cattle-yak was cloned by RT-PCR and analyzed by bioinformatics softwares.qRT-PCR was used to detect the expression patterns of PPP1R11 mRNA in different tissues of cattle-yak,and compared the expression difference in testes at different development stages between cattle-yak and yak.The cell location and expression of PPP1R11 protein were detected by immunohistochemistry(IHC)staining.The results showed that the CDS region of PPP1R11 gene in cattle-yak was 324 bp,which encoded 107 amino acids.The corresponding PPP1R11 protein of cattle-yak had high homology with other mammals.The protein function prediction showed that PPP1R11 could interact with PPP1R2,PPP1R7,PPP1CB,UTP20 and other proteins,and mainly related to the phosphorylation of proteins,which could regulate biological processes such as testicular development,spermatogenesis and sexual maturity.PPP1R11 gene was wildly expressed in various tissues of cattle-yak,and the relative expression level in testis was significantly higher than other tissues(P<0.01).The expression of PPP1R11 gene in cattle-yak testis increased with age.Furthermore,there was significant difference of the expression level of PPP1R11(P<0.01)in testis at juvenile and adult stage of cattle-yak compared with the counterpart of yak.In addition,IHC staining result showed that PPP1R11 protein was significantly lower expressed in cattle-yak spermatogonia and sertoli cells,and there was significant difference compared with yak,and the number of primary spermatocytes in cattle-yak was reduced significantly and meiosis arrested at this stage.This study showed that there were temporal and spatial differences in the expression of PPP1R11 between cattle-yak and yak testis,suggesting that PPP1R11 may be related to to male cattle-yak infertility.However,its specific mechanism needs to be further studied.